Metabolism of acetaldehyde by human erythrocytes.

Acetaldehyde metabolism in human erythrocytes was studied using head-space gas chromatographic determination methods, and it was found that acetaldehyde is metabolized in erythrocytes by NAD dependent cytosolic enzyme having an apparent Km value for acetaldehyde approximately 0.7 mM at pH 7.4, and more than 50% of this activity was reduced by 1 μM disulfiram. So, it is suggested that erythrocytes may have an enzyme system similar to the high Km isozyme of the liver aldehyde dehydrogenase.     

Effect of the presence of hardened erythrocytes on deformation-orientation characteristics of normal erythrocytes in shear flow studied by the spin label method

The effect of the presence of hardened red blood cells (HRBC) in a mixed suspension on the deformation-orientation characteristics of normal cells in flow is experimentally probed by the electron spin resonance (ESR) spin label method, using a phosphatidylcholine label which does not transfer between the cells. The average deformation-orientation of the normal cells is generally suppressed by the presence of HRBC to different degrees, depending upon the shape and the way the HRBC are prepared. The effects are qualitatively explained by disturbance of laminar shear flow due to the random tumbling of the HRBC.     

Reaction of acetaldehyde with hemoglobin

Acetaldehyde reacted with hemoglobin at neutral pH and 37 degrees C to form adducts that were stable to dialysis and that were not reduced by sodium borohydride. Hemoglobin tetramers having 2, 3, and probably 4 molar eq of bound aldehyde were isolated by cation exchange chromatography. The sites of attachment of the aldehyde were the free amino groups of the N-terminal valine residues of the alpha and beta chains of hemoglobin. Derivatization of the beta chains caused a greater increase in the acidity of the hemoglobin than did derivatization of the alpha chains. Derivatization of the beta chains was also preferred over that of the alpha chains. Acetaldehyde derivatives of the N-terminal octapeptide of hemoglobin S (beta sT-1 peptide), Val-Gly-Gly, and tetraglycine were formed readily, contained 1 M eq of acetaldehyde/mol of peptide, and were not reduced by sodium borohydride. In contrast, Ala-Pro-Gly failed to form a 1:1 adduct with acetaldehyde. 13C NMR analysis of the peptide adducts formed with [1,2-13C]acetaldehyde indicated that tetrahedral diastereomeric derivatives were produced. The 13C chemical shifts of the adducts formed between hemoglobin and [1,2-13C]acetaldehyde were identical to those of the peptide adducts although resonances from the individual diastereomeric adducts at each hemoglobin site could not be resolved. The results cited above as well as other evidence indicate that acetaldehyde reacts with the amino termini of hemoglobin to form stable cyclic imidazolidinone derivatives. An exchange of acetaldehyde residues between peptides was also documented.     

Metabolic effects of acetaldehyde

Acetaldehyde, the toxic product of ethanol metabolism in the liver, covalently binds to a variety of proteins, thereby altering liver function and structure. Through its binding to tubulin, acetaldehyde decreases the polymerization of microtubules thereby impairing protein secretion and favouring their retention, with associated swelling of hepatocytes. Acetaldehyde adduct formation also impairs some enzyme activities. Either directly or through binding with GSH, acetaldehyde favours lipid peroxidation. Various mitochondrial functions are altered, particularly after chronic ethanol consumption which sensitizes the mitochondria to the toxic effects of acetaldehyde. In cultured myofibroblasts, acetaldehyde stimulates collagen production. The acetaldehyde-protein adducts stimulate the production of antibodies directed against the acetaldehyde epitope. This immune response may contribute to the aggravation or perpetuation of alcohol-induced liver damage. Some acetaldehyde effects, however, could conceivably be considered as beneficial, such as the stimulation of vascular prostacyclin release which may take part in the 'protective' effect of moderate ethanol consumption against some cardiovascular complications.     

Interaction of erythrocytes with aminoalkylagarose

Some properties of aminoalkyl-agarose as a carrier for adsorption immobilization of erythrocytes were being studied. The effect of the hydrophobic spacer on the erythrocyte affinity for the matrix is shown. The erythrocyte adsorption is characterized by a positive cooperativity; the shorter the spacer, the higher is cooperativity.     

Amplification of antioxidant activity of catechin by polycondensation with acetaldehyde

Catechin exhibits numerous biological and pharmacological effects attributed to antioxidant action. The synthetic poly(catechin)s condensed through acetaldehyde with different molecular weights were assessed in terms of antioxidant activity and enzyme inhibitory activity on the basis of a catechin repeating unit and compared with monomeric catechin. The poly(catechin)s showed great amplification of superoxide scavenging activity, xanthine oxidase (XO) inhibitory activity, and inhibition effects on human low-density lipoprotein oxidation initiated by 2,2'-azobis(2-amidinopropane)dihydrochloride (AAPH) as a radical generator on the catechin unit level, compared to monomeric catechin: these activities were proportional to their molecular weights. The reducing power of the polymer was lower than that of monomeric catechin, which decreased with increasing the molecular weight. The polymer also protected endothelial cells from oxidative injury induced by AAPH, with a greater effect expressed on a catechin unit basis than that of the monomer. These results demonstrate that the poly(catechin)s are more potent antioxidant agents and enzyme inhibitors.     

Liver collagen of rats submitted to chronic intoxication with acetaldehyde

It was found that chronic intoxication of rats with acetaldehyde results in a distinct, progressive increase of 5-³H-proline incorporation into collagen synthesized by liver. At the same time biosynthesis of other proline-containing (noncollagenous) proteins does not change significantly. On the other hand the collagen content in the rat liver did not increase in the early stage of acetaldehyde administration, but increased when acetaldehyde feeding was continued for 6 months. About 40% increase of total collagen content was found in livers of the intoxicated animals. All the investigated collagen types (I, III, IV and V) grew in the same degree. No changes in proportional relationships between collagens of different types were found.     

Overproduction of a stable acetaldehyde dehydrogenase with high affinity to acetaldehyde by Escherichia coli expressing the ACOD gene of Alcaligenes eutrophus

Summary The acoD gene of Alcaligenes eutrophus, which encodes a very stable NAD dependent aldehyde dehydrogenase with high affinity toward acetaldehyde (K _m = 4M), was overexpressed in Escherichia coli. Plasmid pDel087, a deletion derivative of a plasmid constructed recently (Priefert et al., 1992), conferred acetaldehyde dehydrogenase activity of 2.5 U/mg of protein to E. coli, which was about 8-fold higher than the activity in ethanol-grown cells of A. eutrophus.     

A mutagenicity assessment of acetaldehyde

Acetaldehyde has been shown in studies by several different laboratories to be a clastogen (chromosome-breaking) and inducer of sister-chromatid exchanges in cultured mammalian cells (Chinese hamster cells and human lymphocytes). Although there have been very few studies in intact mammals, the available evidence suggests that acetaldehyde produces similar cytogenetic effects in vivo. The production of cytogenetic abnormalities may be related to the ability of acetaldehyde to form DNA-DNA and/or DNA-protein cross-links. Acetaldehyde apparently has not been evaluated for its ability to cause gene mutations in cultured mammalian cells, but it has been shown to produce sex-linked recessive lethals in Drosophila. In general, bacteria tests have been negative. Although acetaldehyde is a genotoxic cross-linking agent, it does not appear to cause DNA strand breaks. There were no studies available regarding the potential of acetaldehyde to produce genetic damage in mammalian germ cells in vivo. Most mutagenicity testing on acetaldehyde has been motivated by attempts to define the proximate mutagen in ethanol metabolism.     

Immobilization of microorganisms with PVA hardened by iterative freezing and thawing

In order to utilize polyvinyl alcohol (PVA) as a gel matrix for the immobilization of microorganisms, PVA was subjected to iterative freezing-thawing. The effects of the procedure on the mechanical characteristics of the PVA hydrogel and the stability of the immobilized microorganisms were investigated. PVA showed rubber-like elasticity after iterative freezing-thawing. Gel strength increased with the iteration number of freezing-thawing until seven iterations. Although the activities of both the free and immobilized cells decreased during the iteration of freezing-thawing, addition of cryoprotectants such as glycerol and skim milk was effective for preventing the decrease in activity.     

The effect of glycosaminoglycans with acetaldehyde on the activation of prothrombin

Heparin17-19k, (25, 50, and 100 ng), heparin6k (50 and 100 ng), heparin3k (50, 100, and 200 microg), chondroitin sulfates B (dermatan sulfate) (0.25, 0.5, and 1.0 microg), C (1 and 10 microg), and A (1 and 10 microg) each prolong the activated partial thromboplastin time (APTT) when preincubated with prothrombin to a greater extent than when preincubated with Factor II-deficient plasma prior to their mixing and subsequent additions of APTT reagent and Ca2+. In all cases statistical significance (p < or = 0.05) was observed except with the 2 lower levels of heparin3k. These results suggest that the glycosaminoglycans (GAGs) may exert a direct effect upon prothrombin (FII) in their anticoagulant activity. Pre mix tures of [(FII/25 ng H17-19k) + 447 mmol acetaldehyde (AcH)/L] as well as [(AcH/H) + FII] and [(FII/AcH) + H] each exert a synergistic anticoagulant effect upon APTT. At low AcH concentrations (44.7 mmol/L), neither a synergistic nor an additive effect is seen. H6k and H3k, on premixing with 447 mmol AcH/L, exhibit an additive effect on APTT prolongation but no synergism. Similarly, premixtures of CSB/447 mmol AcH/L/FII show a greater anticoagulant effect than do [(CSB/AcH) + FII] or [(FII/AcH) + CSB] premixtures. CSC-AcH and CSA-AcH patterns are analogous to those of CSB (DS). These data suggest the possibility that AcH, the primary product of ethanol metabolism, may serve as a crosslinking adduct with proteins, in this case, prothrombin, as well as GAGs. Thus ternary complexes between the zymogen form of coagulation factors, GAGs, and AcH are possible, further influencing coagulopathy.     

The interaction of phenyldichloroarsine with erythrocytes

The purpose of the study was to identify binding sites of organic arsenic in the erythrocyte and to explain species differences in binding. Washed erythrocytes were exposed to graded concentrations of [U-14C]phenyldichloroarsine (PDA) in phosphate-buffered saline containing 0.1% glucose and 0.1% bovine serum albumin. At low PDA concentrations, all cells bound the arsenical rapidly (within 10 min) and quantitatively. Human, pig, hamster, guinea pig, and mouse erythrocytes approached saturation at 0.02-0.3 mumol PDA/10(9) cells, depending on the species. Saturation points correlated well with each respective species' erythrocyte glutathione content. In contrast, rat erythrocytes showed no sign of saturation at PDA loads as high as 3.0 mumol/10(9) cells. Hemolysates of PDA-treated erythrocytes were subjected to Sephadex G-75 gel filtration chromatography. 14C from rat hemolysate was distributed between the hemoglobin and small molecular weight (glutathione-containing) fractions. In all other species, the 14C eluted almost exclusively with the glutathione-containing fractions. In equilibrium dialysis experiments, human hemoglobin did not bind PDA, whereas rat hemoglobin bound 2 PDA/mol with Kd approximately 5 microM. In conclusion, glutathione is the principal binding site of phenyldichloroarsine in erythrocytes. In most species, the arsenical does not bind to hemoglobin, even though it has free (titratable) sulfhydryls considerably in excess of the glutathione concentration. In rat erythrocytes, phenlydichloroarsine binds both to glutathione and to hemoglobin. Arsenical binding by rat hemoglobin is presumably due to the unique location of the extra titratable cysteine in that protein.